title: Purification and biochemical characterization of glutathione S-transferase from Down syndrome and normal children erythrocytes: a comparative study.
abstract: Down syndrome (DS) is the phenotypic manifestation of trisomy 21. Our study was concerned with the characterization and purification of glutathione S-transferase enzyme (GST) from normal and Down syndrome (DS) erythrocytes to illustrate the difference in the role of this enzyme in the cell. Glutathione S-transferase and glutathione (GSH) was determined in ten DS and ten healthy children matched for age (3-10 years). DS group exhibited significantly lower GST value (2.7 units/gHb) as compared to controls (6.6 units/gHb) (40.9%). GST activity was significantly decreased to 40.9% in the DS group as compared to controls. Also GSH concentration was significantly decreased to 60.6% in the DS group compared to the controls. Glutathione transferase was purified from erythrocytes of normal and DS pooled blood samples by affinity chromatography with specific activity of 23.7% and 7.9%, respectively. The effect of freezing and thawing, storage time of freezing and GSH concentration on the stability of the enzyme were examined. Normal GST exhibited a pH optimum at pH 7 followed by sharp decrease, however DS GST exhibited pH optimum between pH 7.5 and 8. The Km values for 1-chloro-2,4-dinitrobenzene (CDNB) and GSH were 0.205 mM and 0.786 mM, respectively, for normal GST, and 0.318 mM and 1.307 mM, respectively for DS GST. The activation energy (Ea) was calculated to be 2.25 and 4.25 cal/mol for normal GST and 3.8 cal/mol for DS GST. Normal and DS GST were inhibited by the same inhibitors (hematin, bromosulfophthalein and cibacron blue), but with different degree. On kinetic basis, the individuals with lower overall GST activity and slight differences in some kinetic characters are at greater risk from xenobiotic contamination as compared to those with higher overall GST activity observed in normal individuals.
authors.affiliation: Department of Molecular Biology, National Research Center, Dokki, Cairo, Egypt.
authors.forename: Ragaa R
authors.initials: RR
authors.lastname: Hamed
authors.forename: Tahany M
authors.initials: TM
authors.lastname: Maharem
authors.forename: Nagwa
authors.initials: N
authors.lastname: Abdel-Meguid
authors.forename: Gilane M
authors.initials: GM
authors.lastname: Sabry
authors.forename: Abdel-Monem
authors.initials: AM
authors.lastname: Abdalla
authors.forename: Rasha A
authors.initials: RA
authors.lastname: Guneidy
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D002384:Catalysis; D002648:Child; D002675:Child, Preschool; D002846:Chromatography, Affinity; D004314:Down Syndrome / Q000201:enzymology*; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004912:Erythrocytes / Q000201:enzymology*; D005978:Glutathione / Q000378:metabolism; D051549:Glutathione S-Transferase pi / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007223:Infant; D018384:Oxidative Stress / Q000502:physiology*; D013379:Substrate Specificity; D013694:Temperament
chemical_list: D000975:Antioxidants; D004791:Enzyme Inhibitors; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005978:Glutathione